In previous work we have demonstrated that Heat Shock Factor 1 (HSF1), master regulator of the mammalian heat shock response acts in a powerful, multi-faceted manner to enable malignant transformation both in cell culture and in transgenic mice. When compared to Hsf1 wild type mice, transgenic mice in which Hsf1 is knocked out are dramatically resistant to the formation of cancers driven by a variety of oncogenic lesions including activating mutations of RAS and hot-spot mutations of the tumor suppressor p53. In addition to highlighting the antitumor activity of targeting HSF1, the ability o mice to tolerate complete genetic knockout of HSF1 suggests that a useful therapeutic index is likely to exist for selective inhibitors of HSF1. Unfortunately, no such small molecules are currently known. Therefore, the goal of this project is to develop drug-like inhibitors of the HSF1 regulated transcriptional program with potent and selective activity in mice. We hypothesize that such inhibitors will be invaluable in probing how this ancient, highly conserved stress response makes it possible for cells to cope with the problems imposed by malignancy. They will also serve as promising leads for the future development of useful anticancer drugs with a completely new mode of action. Supported by a previous R-03 grant to one of the principal investigators on this project, we recently completed a >300,000 compound high throughput screen (HTS) through the NIH MLPCN program designed to identify selective inhibitors of HSF1. One of our most promising hits was the natural product rocaglamide A (RocA). To move from screen hit to a probe with potent and selective activity in animals, a medicinal chemistry collaboration has been established with another investigator who leads a nearby Chemical Methodology and Library Development (CMLD-BU) center. His group has achieved directed synthesis of diverse structural analogs of RocA starting from simple materials. Our joint expertise and resources ensure an adequate supply of material for biological studies independent of limited natural resources and have allowed us to pursue initial structure activity relationship (SAR) studies. These have identified compounds with more potent anticancer activity, but significant target-specificity issues and pharmacological liabilities remain to be addressed. In direct response to the mandate of PAR-12-060, we propose to undertake iterative bioassay and chemical optimization cycles to transition a validated HTS hit to a useful in vivo chemical probe.